In order to purify nucleic acid from a sample containing the nucleic acid, physical and/or chemical purification processes need to be carried out. In most cases, it is general to remove a large-sized solid component from a liquid in the process of purifying the nucleic acid.
In particular, in the case where the nucleic acid is obtained by dissolving a biological sample, it is necessary to remove impurities including various kinds of protein components contained in a cell. In general, the impurities are removed by a precipitation method using centrifugation or by a filtering method with a filter. The former is frequently used in a laboratory, but is not appropriate in treating a large amount of samples and difficult to automate. Whereas, the latter is appropriate in continuously treating a large amount of samples by being applied to an automated device with a pressure reducing or applying device.
Chromosomal DNA needs to be isolated in the process of extracting only plasmid from Escherichia coil or bacterial transformants since an undesired reaction due to the chromosomal DNA needs to be prevented to progress during a gene amplification process, a gene sequencing process, a gene recombination process, or the like.
A method of separating nucleic acid from a tissue sample is largely divided into two steps. As the first step, tissue cells are lysed to thereby release all the cell components including all nucleic acids. This cell lysis procedure may be conducted by a physical or chemical method. However, the chemical method is widely used in order to minimize damage of the nucleic acid and effectively lyse tissue cells to thereby isolate the nucleic acid. In particular, an anionic surfactant such as sodium dodecyl sulfate (SDS) is mainly used.
As the second step, impurities are removed from the cell lysate to isolate pure nucleic acid. This method of extracting the nucleic acid uses treatment of an organic solvent such as phenol-chloroform, salting out, a chaotropic salt, and the like.
The method of separating nucleic acid from tissue cells through the above two steps is widely used, and thus, according to the existing methods of isolating nucleic acid, a cell lysis buffer containing a surfactant is used as a first buffer for extracting nucleic acid in order to facilitate isolation of nucleic acid from tissue cells. A second buffer in the process of separating nucleic acid is a buffer containing a high-concentration salt such as a chaotropic salt. The buffer containing this high-concentration salt has a very important role in removing impurities from the nucleic acid by salting out.
In order to isolate high-purity nucleic acid, the two kinds of buffers as described above are applied step by step to the tissue cells to thereby be used to remove impurities through cell lysis and salting out. Particularly, a cell lysate lysed in the cell lysis buffer containing sodium dodecyl sulfate is mixed with a buffer containing a high-concentration salt, followed by salting out, so that precipitation of impurities can be facilitated, and thus, this is very important in separating high-purity nucleic acid, and most widely used as a cell lysis buffer for effective cell lysis in the existing method of isolating nucleic acid.
Recently known nucleic acid isolation techniques are used in isolating the nucleic acid from the cell lysate by using a silica membrane or ion exchange chromatography. Particularly, according to the current widely used method, nucleic acid is isolated from the other impurities by being bound on a silica membrane, and the bound nucleic acid is obtained through washing and elution [Process for isolating nucleic acid (U.S. Pat. No. 5,234,809)].
However, in this method, the nucleic acid may be isolated from the impurities by being bound on the silica membrane, but a centrifugation step is necessary in all the steps of isolating the nucleic acid, and thus, the nucleic acid isolation procedure is difficult and needs to be performed only at a laboratory.
Moreover, when the two kinds of buffers as described above are mixed, they are severely crystallized, and thus may not be used in a mixture type, and the cell lysate lysed using this type of buffers may not be applied to a silica membrane, and thus, is not very appropriate as a high-speed nucleic acid solution.
Meanwhile, according to the nucleic acid isolation technique using the ion exchange chromatography, negatively charged nucleic acid is combined with a cation exchange resin and then isolated from impurities. This method necessarily requires isolation of nucleic acid using centrifugal separation, and is not appropriate in isolating nucleic acid for point-of-care testing (POCT) due to the need of a skilled person and complicated steps.
Also, in a case of a method of binding nucleic acid contained in a sample to magnetic particles combined with a carboxyl group and then isolating the nucleic acid from impurities, a large amount of samples need to be used, and this method is not appropriate in isolating nucleic acid for point-of-care testing (POCT) due to a complicated separation step of using the magnetic particles and a complicated nucleic acid separation step of separating the magnetic particles combined with the nucleic acid using centrifugation or magnet.